The Research Of Antigenic Peptides Covalently Linked With Immunoadjuvant-pulsed Dendritic Cells As A Cancer Vaccine

It is well known that,cancer is the second leading cause of death after heart diseases.Even though,the standard treatment strategies of cancer（such as chemotherapy and radiotherapy）can improve patients’survival rate,but they are often accompanied by drug resistance and side effects.In recent years,immunotherapy has attracted great attention due to its promising preliminary results in achieving meaningful and durable treatments responses with minimal manageable toxicity.Among them,therapeutic tumor vaccines can reduce tumor volume or provide patients with protection against recurrence.Dendritic cells are the most powerful professional antigen presenting cells,mediating innate immunity and inducing adaptive immunity.Thus,they are considered as an ideal antigen delivery vector for cancer vaccines.Several studies showed that peptides are easy to synthesis,stable to store,safe and can induce specific immune responses.Thus,peptide-based DC vaccine hold promise as effective strategy to treat cancer.However,the success of this strategy mainly depends on antigenic peptides and the selection of an appropriate adjuvant.The aim of this study was to elicit a broad immune response against multiple epitopes with the assistance of the immunoadjuvant HB_100-108（HMGB1-derived peptide）.Three HLA-A*0201 restricted peptides（Survivin,Her2 and CEA）were synthesized and covalently linked with HB_100-108via a double arginine linker（RR）,followed by loading them on human monocyte-derived DCs.Fluorescence microscopy and flow cytometry were used to detect the uptake efficacy of peptides by DC.The maturation and activation status of peptides-pulsed DC was evaluated by detecting the expression of DC surface molecules and the secretion level of cytokines.The ability of peptides-pulsed DC to activate T cells was evaluated by degranulation assay and detection of secreted cytokines.The lactate dehydrogenase assay was used to evaluate the cytotoxicity of effector T cells against tumor cells in vitro.Results showed that,DC could efficiently take up the antigenic peptides covalently linked with HB_100-108.Furthermore,Compared with control group,DCs/Peptide group and DCs/HB_100-108group,DCs/Peptide-HB_100-108group expressed higher levels of MHC class I and class II molecules（HLA-ABC and HLA-DR）,costimulatory molecules（CD80,CD86 and CD40）,maturation marker CD83 and chemokine receptor 7（CCR7）,and produced more proinflammatory cytokines（TNF-α,IL-6,IFN-γand IL-12）.Moreover,DC loaded with peptide-HB_100-108efficiently activated T cells through enhancing the secretion of inflammatory cytokines.Besides,DC loaded with peptide-HB_100-108induced a potent degranulation of T cells as identified by the expression of surface marker CD107a,indicating that those T cells could efficiently secrete cytotoxic effector molecules against their targets.Finally,Peptide-HB_100-108greatly enhanced the cytotoxicity of T cells against pancreatic cancer cell line（PANC-1）in vitro.In conclusion,these results indicate that antigenic peptides covalently linked with HB_100-108-pulsed DCs could be an efficient strategy for improving DC-based tumor vaccines.